Differential regulation of STP, LTP and LTD by structurally diverse NMDA receptor subunit-specific positive allosteric modulators.

Different types of memory are thought to rely on different types of synaptic plasticity, many of which depend on the activation of the N-Methyl-D Aspartate (NMDA) subtype of glutamate receptors. Accordingly, there is considerable interest in the possibility of using positive allosteric modulators (PAMs) of NMDA receptors (NMDARs) as cognitive enhancers. Here we firstly review the evidence that NMDA receptor-dependent forms of synaptic plasticity: short-term potentiation (STP), long-term potentiation (LTP) and long-term depression (LTD) can be pharmacologically differentiated by using NMDAR ligands. These observations suggest that PAMs of NMDAR function, depending on their subtype selectivity, might differentially regulate STP, LTP and LTD. To test this hypothesis, we secondly performed experiments in rodent hippocampal slices with UBP714 (a GluN2A/2B preferring PAM), CIQ (a GluN2C/D selective PAM) and UBP709 (a pan-PAM that potentiates all GluN2 subunits). We report here, for the first time, that: (i) UBP714 potentiates sub-maximal LTP and reduces LTD; (ii) CIQ potentiates STP without affecting LTP; (iii) UBP709 enhances LTD and decreases LTP. We conclude that PAMs can differentially regulate distinct forms of NMDAR-dependent synaptic plasticity due to their subtype selectivity.

Initiating and imaging the coherent surface dynamics of charge carriers in real space.

The tip of a scanning tunnelling microscope is an atomic-scale source of electrons and holes. As the injected charge spreads out, it can induce adsorbed molecules to react. By comparing large-scale 'before' and 'after' images of an adsorbate covered surface, the spatial extent of the nonlocal manipulation is revealed. Here, we measure the nonlocal manipulation of toluene molecules on the Si(111)-7 × 7 surface at room temperature. Both the range and probability of nonlocal manipulation have a voltage dependence. A region within 5-15 nm of the injection site shows a marked reduction in manipulation. We propose that this region marks the extent of the initial coherent (that is, ballistic) time-dependent evolution of the injected charge carrier. Using scanning tunnelling spectroscopy, we develop a model of this time-dependent expansion of the initially localized hole wavepacket within a particular surface state and deduce a quantum coherence (ballistic) lifetime of ∼10 fs.

Spectroscopic measurements in scleritis: bluish-red or deep red?

PURPOSE: To design a slit-lamp mountable spectrometer for the assessment of ophthalmic patients and to illustrate a potential clinical application by measuring the spectral characteristics of inflamed eyes of differing aetiologies. METHODS: A slit lamp mountable instrument was designed and built, and methods for data analysis developed. Reflectance spectra were recorded from two patients with scleritis, three with non-scleritic red eyes and from two controls with non-inflamed eyes. RESULTS: Measurements were reproducible and demonstrated statistically significant differences in the spectral characteristics between the three groups. Spectra from scleritic eyes revealed a relative increase in intensity of long wavelength red light, 650-740 nm, compared with non-scleritic red eyes. These longer wavelengths will be appreciated as dark red. There was no increase in relative intensity in the blue part of the spectrum in scleritic eyes. CONCLUSIONS: Reproducible measurements of the eye were made using an innovative, slit-lamp mountable spectrometer and its functionality demonstrated by differentiating the spectra from eyes with differing pathologies. While intending only to illustrate one potential application; for the cases examined, our results indicate that inflamed scleritic eyes exhibit a longer wavelength red light with no increase in shorter wavelength blue light. Thus our measurements would seem to confirm that the perceived redness of scleritis differs from other red eyes. However, it is a deeper darker red and not a bluish one as traditionally described.

Excitatory inputs to spiny cells in layers 4 and 6 of cat striate cortex.

The principal target of lateral geniculate nucleus in the cat visual cortex is the stellate neurons of layer 4. In previously reported work with intracellular recording and extracellular stimulation in slices of visual cortex, three general classes of fast excitatory synaptic potentials (EPSPs) in layer 4a spiny stellate neurons were identified. One of these classes, characterized by large and relatively invariant amplitudes (mean 1.7 mV, average coefficient of variation (CV) 0.083) were attributed to the action of geniculate axons because, unlike the other two classes, they could not be matched by intracortical inputs, using paired recording. We have examined in detail the properties of this synaptic input in twelve examples, selecting for study those EPSPs where there was secure extracellular stimulation of the single fibre input to a pair of stimuli 50 ms apart. In our analysis, we conclude that the depression that these inputs show to the second stimulus is entirely postsynaptic, since the evidence strongly suggests that the probability of transmitter release at the synaptic site(s) remains 1.0 for both stimuli. We argue that the most plausible explanation for this postsynaptic depression is a reduction in the average probability of opening the synaptic channels. Using a simple biochemical analysis (c.f. Sigworth plot), it is then possible to calculate the number of synaptic channels and their probability of opening, for each of the 12 connections. The EPSPs had a mean amplitude of 1.91 mV (+/- 1.3 mV SD) and a mean CV of 0.067 (+/- 0.022). The calculated number of channels ranged from 20 to 158 (59.4 +/- 48.7) and their probability of opening to the first EPSP had an average of 0.83 (+/- 0.09), with an average depression of the probability to 0.60 for the second EPSP. Geniculate afferents also terminate in layer 6. Intracellular recordings were also made in the upper part of this layer and a total of 51 EPSPs were recorded from pyramidal cells of three principal types. Amongst this dataset we sought EPSPs with similar properties to those characterized in layer 4a. Three examples were found, which is a much lower percentage (6%) than the incidence of putative geniculate EPSPs found in layer 4a (42%).

Excitatory synaptic inputs to spiny stellate cells in cat visual cortex.

In layer 4 of cat visual cortex, the monocular, concentric receptive fields of thalamic neurons, which relay retinal input to the cortex, are transformed into 'simple' cortical receptive fields that are binocular and selective for the precise orientation, direction of motion, and size of the visual stimulus. These properties are thought to arise from the pattern of connections from thalamic neurons, although anatomical studies show that most excitatory inputs to layer 4 simple cells are from recurrently connected circuits of cortical neurons. We examined single fibre inputs to spiny stellate neurons. We examined single fibre inputs to spiny stellate neurons in slices of cat visual cortex, and conclude that thalamocortical synapses are powerful and the responses they evoke are unusually invariant for central synapses. However, the responses to intracortical inputs, although less invariant, are strong enough to provide most of the excitation to simple cells in vivo. Our results suggest that the recurrent excitatory circuits of cortex may amplify the initial feedforward thalamic signal, subserving dynamic modifications of the functional properties of cortical neurons.

Dendritic morphology of CA1 pyramidal neurones from the rat hippocampus: I. Branching patterns.

The aim of this study was to provide quantitative descriptions of the dendritic branching patterns of pyramidal neurones in the CA1 region of the rat hippocampus. Thirteen adult cells were filled with biocytin and reconstructed by using the light microscope. The number of basal trees arising from the soma of each cell ranged from two to eight. There was wide variation in the number of terminal segments per tree. Six cells had single apical trunks, and seven had trunks that bifurcated in stratum radiatum. The number of apical oblique trees ranged from nine to 30, with each tree usually showing a lower degree of branching than basal trees. Basal and oblique trees had similar branching patterns, with the majority of branch points occurring close to the origin of the tree. Both basal and oblique terminal segments were generally much longer than intermediate segments and constituted up to 90% of the combined dendritic length of the tree. The branching pattern of the apical tuft was different, with many relatively long intermediate segments; terminal segments contributed only some 66% of the combined dendritic length of these trees. The mean total combined dendritic length for six adult cells reconstructed and measured completely was 11,900 +/- 1,000 microns (standard deviation). The relative proportions of the different parts of the dendritic system, although not the total dendritic length, were correlated with the location of the soma relative to the cell body layer. Cells with somata close to the stratum pyramidale/stratum radiatum border had more dendrites terminating in stratum radiatum and fewer in stratum oriens than cells with somata further from it.

Dendritic morphology of CA1 pyramidal neurones from the rat hippocampus: II. Spine distributions.

The numbers and distributions of dendritic spines were estimated for six adult and three juvenile biocytin-injected neurones from the CA1 region of the hippocampus of the albino rat. For each cell, a sample of long dendritic segments that lay favourably in the plane of focus was drawn at high magnification and the visible spines counted. Correction was made for spines obscured by dendritic shafts. Within individual cells, dendrites of similar type and diameter had similar spine densities. For adults, long basal segments averaged 2.4 spines/microns and obliques averaged 3.2 spines/microns. In juveniles, basals averaged 2.3 spines/microns and obliques, 2.5 spines/microns. Apical tuft segments were less spiny, averaging 1.4 spines/microns in adult cells and 1.8 spines/microns in juveniles. There was a positive correlation between spine density and dendrite diameter. Values from this sample were used to assign spine densities to the other segments, and so the total number of spines was estimated for each cell. Adult cells averaged 30,500 +/- 3,900 (S.D.) spines and juveniles, 23,800 +/- 7,100 spines. Adult cells had roughly 50% of their spines in stratum radiatum, 40% in s. oriens, and 10% in s. lacunosum-moleculare. Juvenile cells had a rather higher proportion (20%) in s. lacunosum-moleculare. In general, some 50% of all spines were located within a path length of 200 microns from the soma. These total numbers of spines were much higher than earlier values from Golgi-impregnated cells but align well with estimates of the numbers of axonal boutons supplied to CA1 by CA3 pyramidal cells.

Interacting effects of Ca2+ and hypoxia in the induction of sarcolemmal damage in mouse diaphragm in vitro.

Experiments have been undertaken to investigate the basis for the selective damage of centrally placed fibres in mouse diaphragms exposed to Ca2+ loading in vitro. Incubation under hypoxic conditions (non-aerated saline) for 30 min had no discernible effect on the muscle. Incubation for 120 min led to permeabilisation of the sarcolemma (assayed by penetration of Procion Yellow) in 54% of cells. Sarcolemmal permeabilisation was almost completely restricted to centrally placed cells, as has previously been described for the effects of the Ca2+ channel agonist Bay K 8644. Ultrastructural damage to the myofibrils and mitochondrial swelling were also widespread amongst centrally placed cells. Permeabilisation was inhibited when hypoxic incubations were carried out in Ca(2+)-free saline. Incubation in hypoxic Ca(2+)-containing saline for 30 min followed by further incubation in Ca(2+)-free, hypoxic saline, up to a total of 120 min, resulted in permeabilisation similar to that seen in muscles incubated for 120 min in Ca(2+)-containing, hypoxic saline. It is suggested that selective damage to central fibres induced by Ca2+ loading resulting from treatment with Bay K 8644 is related to increased oxygen demand.

Sodium is elevated in mdx muscles: ionic interactions in dystrophic cells.

Recent evidence suggests that cellular sodium regulation may be abnormal in muscular dystrophy. We have measured intracellular sodium concentration (Nai) in muscles of mdx mice (a model of Duchenne muscular dystrophy) using two techniques. Nai in isolated diaphragm (measured using a microelectrode) was 13.0 +/- 0.3 mM and 23.5 +/- 0.7 mM (mean +/- SE) in the control and mdx mice respectively. Nai in gastrocnemius muscle (calculated from extra- and intracellular volumes using serum and whole-muscle sodium concentrations) was 13 +/- 3 mM and 24 +/- 2 mM (mean +/- SE) in control and mdx, respectively. We argue that this abnormality in mdx tissues could reflect a reduced flux through the Na/K ATPase, although a contribution from increased Na leak cannot be ruled out. We also discuss possible consequences of an increased Nai: for example, raised Nai may lead to defective cell volume control in Duchenne dystrophy and the mdx mouse.

Immediate and delayed effects of a brief period of calcium loading, induced by the calcium channel agonist Bayer K 8644, on the ultrastructure and sarcolemmal integrity of murine diaphragm in vitro.

Exposure of in vitro mouse diaphragm to the dihydropyridine Ca2+ channel agonist Bayer K 8644 (Bay) and partial depolarisation for 120 min induced severe structural damage within 30-60 min and membrane permeabilisation (60-120 min). Exposure to Bay and depolarisation for 30 min (brief Ca2+ loading) followed by washout for 90 min produced similar effects, even though significant membrane damage does not occur until the second hour of incubation. Development/expression of necrosis during the washout period was not inhibited by manoeuvres designed to combat the effects of any remaining Bay, or by use of a Ca(2+)-free, 1 mM EGTA medium for washout. Exposure to Bay for 15 min followed by washout did not induce damage. Electron microscopic examination revealed that the mitochondria of cells fixed after 30 or 120 min exposure to Bay and depolarisation were in the orthodox (swollen) configuration, but that mitochondria in muscles that had undergone washout with Ca(2+)-free-EGTA saline were electron dense. Further incubation of these muscles in Ca(2+)-containing medium caused readoption of the swollen configuration. We conclude that membrane permeabilisation can occur after removal of the Ca2+ load, and that mitochondrial Ca2+ overload is not necessary for Ca(2+)-induced damage/death to occur.